FORMULATING FOODS TO
CONTROL BACTERIAL
PATHOGENS
Kathleen Glass, Ph.D.
1550 Linden Drive, Madison, WI 53706
Email: [email protected]
Wisconsin Association for Food Protection Food Safety Workshop
June 13, 2017, Madison, WI
Agenda
• Background
• Food safety risks
• Critical factors controlling microbial growth
• Available moisture (water activity)
• Acidity (pH)
• Ingredients with antimicrobial activity
• Validating formulation as a preventive control in
food safety plan
• Examples of controlling bacterial pathogens
Microbes Cause Foodborne Illness
• 48 Million episodes of foodborne illness per year
• Viruses, bacteria, parasites
• 128,000 hospitalizations
• Salmonella, Campylobacter, Norovirus
• 3,000 deaths
• Salmonella, Listeria monocytogenes
• Toxoplasma gondii
• Economic impact
• Direct medical costs & lost wages
• Recall costs and litigation/liability• Peanut butter – Salmonella 2008-2009, Recall cost $50-60 million;
Peanut Corp. Am. bankrupt
Scallon et al., 2011; University of Florida Emerging Pathogens Institute, 2011
Factors that contribute to foodborne
illness (typically more than one)Contamination of raw
commodities
Flour, E. coli O121 and O26
Soynut butter, E. coli O157:H7
Raw milk gouda, E. coli O157:H7
Sprouts, Salmonella
Slow acid development Staphylococcus aureus cheese, yogurt,
fermented sausage
Improper hot-holding temperature Clostridium perfringens, buffets, catering
Clostridium botulinum, nacho cheese sauce
Recontamination of products Listeria monocytogenes, ice cream
Ability of microbes to grow at
refrigeration temperatures
Listeria monocytogenes, packaged salads
Lack of growth inhibitors Listeria monocytogenes, soft, high pH
cheeses
Temperature abuse Clostridium botulinum, carrot juice
Susceptible Consumers Listeria monocytogenes, cantaloupe
5
Bacterial Pathogens: General Concepts
• Pathogens generally found at low levels
• Pathogens do not always cause spoilage
• Can survive or grow in adverse conditions
• Survive cooking; grow under refrigeration temperatures
• Infectious dose varies
• Toxin formation requires growth
• No growth = no toxin = no illness
• Examples: S. aureus, B. cereus, C. botulinum
• Growth can occur in hours, days or weeks
Formulation Strategies Focus on
Gram Positive Bacteria• Tend to be more resistant to thermal
inactivation (especially spores)
• Typically require growth to cause
illness
• Vegetative pathogens
• Listeria monocytogenes
• Staphylococcus aureus
• Sporeforming bacteria
• Bacillus cereus
• Clostridium perfringens
• Clostridium botulinum
6
Do not rely on temperature alone
• Cooking/pasteurization is not perfect
• Spores survive heating
• Post-pasteurization contamination
• Temperature abuse is common
• During distribution, transportation,
consumer homes, power-outages
• Growth of psychrotrophic pathogens
• Listeria monocytogenes
• Some Clostridium botulinum strains
• Some Bacillus cereus strains
Formulating Foods for Safety
• Goal: Maintain safety through the point of
consumption
• Delay pathogen growth until gross spoilage
• Requires understanding of microbial physiology
and ecology in foods
• Adequacy of control depends on target microbe
• Consider whole food, individual components, and
interfaces of components
Critical Factors for Formulation
Available moisture • Water activity aw
• Function of moisture, salt, other ingredients
pH• Acid type, titratable acidity
Addition of growth inhibitors• Preservatives (synthetic or clean label/”natural”)
• Additive or synergistic interaction means that lower levels of each factor can be used
o Less effect on sensory attributes
Use in combination with temperature control
9
pH and aw combinations that inhibit
growth of vegetative cells and spores
Critical aW
values
Critical pH values
<4.2 4.2 – 4.6 >4.6 – 5.0 >5.0
<0.88 No growth No growth No growth No growth
0.88 – 0.90 No growth No growth No growth ?
>0.90 – 0.92 No growth No growth ? ?
>0.92 No growth ? ? ?
? = Requires time/temperature control unless product testing demonstrates otherwise
Adapted from: IFT. 2001. Evaluation and Definition of Potentially Hazardous Foods, IFT/FDA Contract No. 223-98-2333.
Antibacterial ingredients
Synthetic Clean Label
Lactate*, propionate* Cultured sugar, cultured milk
Diacetate*, acetic acid* Dry vinegar, buffered vinegar
Nitrite* Cultured celery
Erythorbate, ascorbate* Acerola cherry powder
Sorbic acid None (derived from rowanberries)
Benzoic acid Cranberries, prunes, plums, cinnamon
Phenolics, flavonoids Fruit / spice extracts
* Clean label substitute with documented efficacy
• Secondary barrier during temperature abuse or mishandling
• Typically bacteriostatic (delays growth, does not kill)
Limitations of Preservatives
• NOT a substitute for good
manufacturing practices
• Component of food safety plan:
preventive controls
• Consumer acceptance, effect on
sensory, functional attributes, cost
• Efficacy affected by food components
• Fat level (solubility), moisture,
temperature, pH, nitrite, smoke,
• Competitive microflora may also be
inhibited
Validating Formulation Safety
• Validation establishes the scientific basis for
process preventive controls in the Food Safety
Plan
• Ex. Validate critical limit values for process controls
• May include:
• Using scientific principles and data
• Use of expert opinion (including predictive models)
• Challenging the process at the limits of its operating
controls
• Performed or overseen by a preventive controls
qualified individual (PCQI)
Assessing Risks
• Specific for a given food/characteristics
• Requires understanding of:
• Likelihood of contamination
• Processing and handling in facility
• Distribution
• Shelf-life
• Quality based on microbial changes vs. organoleptic
changes
• Temperature of storage
• Potential use/abuse at retail, consumer level
VALIDATING
FORMULATION SAFETY
EXAMPLESEffect of formulation on processed meat
Listeria monocytogenes (extended refrigerated storage)
Clostridium perfringens (cooling)
Effect of formulation on natural cheese
Listeria monocytogenes
Effect of formulation on process cheese
Staphylococcus aureus
Clostridium botulinum
Effect of formulation on refrigerated meals
Clostridium botulinum
High moisture RTE Meats
L. monocytogenes, 41°F (5C), no growth inhibitors
0
1
2
3
4
5
6
7
8
0 2 4 6
log
cfu
/g
Week
Turkey
Ham
Wieners
Beef
Salami
Glass and Doyle, AEM, 1989
Critical factors
• Competitive
microflora
• pH, moisture
• Nitrite
16
Buffered vinegar, moisture, pH
Uncured RTE Meats: L. monocytogenes
17
-1
0
1
2
3
4
5
6
0 2 4 6Ch
an
ge P
op
ula
tio
ns L
. m
on
ocyto
gen
es (
Lo
g C
FU
/g)
Weeks storage at 40F (4C)
Turkey Control No Antimicrobials
Beef Control No Antimicrobials
Turkey - 2.0% buffered vinegar
Beef - 2.0% buffered vinegar
Turkey: ~73% moisture, pH 6.15, 1.1% NaCl, no nitrite
Beef: ~66% moisture, pH 5.75, 0.6% NaCl, no nitrite
Adapted from JFP 76:1366, 2013
Similar results with 1.5% lemon/cherry/vinegar blend; 3.0% cultured sugar-vinegar blend
Effect of nitrite source + cure accelerator, Clostridium perfringens, 15 h biphasic cooling, cured meats,
Appendix B
-2
-1
0
1
2
3
4
5
6
0 2.5 5 7.5 10 12.5 15
Cha
ng
e log
Cooling time (h)
Uncured
547 ppm purified ascorbate
100 ppm purified nitrite
100 ppm natural nitrite
100 ppm purified nitrite+547 ppm purified ascorbate
100 ppm natural nitrite+547 ppm natural ascorbate
King, et a., J. Food Prot. 78: 1527-1535.
• Deli turkey model system• 74% moisture, 1.3% NaCl, pH 6.3
• No effect of conventional
versus alternative (natural)
ingredients
• 100 ppm nitrite alone not
sufficient
• Requires ascorbate for
inhibition
ComBase Perfringens Predictor
Effect of pH and NaCl, cured meat (NaNO2)
No erythorbate in model, Clostridium perfringens
Similar trends in ham with 156 ppm nitrite plus 547 erythorbate cooled over 25 h
0
1
2
3
4
5
0 5 10 15
Lo
g c
ha
nge
(gro
wth
)
Hours cooling from 140 to 45F
pH 6.4, 1.5% NaCl
pH 6.1, 1.5% NaCl
pH 6.4, 3.0% NaCl
pH 6.1, 3.0% NaCl
Effect of cultured dairy solids (clean label antimicrobial)
on L. monocytogenes on Reduced Salt Mozzarella
Same trends, but less differences (more growth) at pH 6.0
2
3
4
5
6
7
8
0 30 60 90 120
log C
FU
/g
Days at 4°C (40°F)
40F Storage, pH 5.8
pH 5.8, 1.8% NaCl
pH 5.8, 1.0% NaCl
pH, 5.8, 1.0% NaCl, 1% Ferm
2
3
4
5
6
7
8
0 30 60 90 120
log C
FU
/g
Days at 7°C (45°F)
45F Storage, pH 5.8
pH 5.8, 1.8% NaCl
pH 5.8, 1.0% NaCl
pH 5.8, 1.0% NaCl, 1% Ferm
Effect of acid type, pH, and
moisture on L. monocytogenes
Lactic Acid Acetic Acid
22
Process cheese slices, 85°F
FRI, Unpublished data, 2001Sponsored by DMI
0
1
2
3
4
5
6
7
8
0 0.5 1 1.5 2 3 4 7 10 14
Hour s
log
cfu
/g
Minimum detection limit
Population at which
staphylococcal enterotoxin is
detected
Average
S. aureus
Upper SD of
growth for S.
aureus
Results: no growth L. monocytogenesE. coli O157:H7 SalmonellaB. cereus
Variable growth StaphGrowth at 1-7 days;pH>5.6, no sorbate
40% moisture for Cheddar based
20 formulations tested; Staph greatest variation for growth
23
S. aureus on process cheese and cheese food
Effect of pH and water activity/moisture
FRI, unpublished data, 2004; sponsored by industry
Pepper Jack (45%moisture, pH, 5.5,
1.7% NaCl, aw0.945)
American (43%moisture, pH 5.3,1.8% NaCl, aw
0.955)
Cheddar2 (42%moisture, pH 5.3,2.0% NaCl, aw
0.955)
Cheddar1 (42%moisture, pH 5.6,1.8% NaCl, aw
0.945)
Cheddar3 (40%moisture, pH 5.3,2.5% NaCl, aw
0.935)
0
1
2
3
1
2
4
Wee
ks a
t 8
0F
Lo
g c
ha
nge
Pepper Jack (45% moisture, pH, 5.5, 1.7% NaCl, aw 0.945)
American (43% moisture, pH 5.3, 1.8% NaCl, aw 0.955)
Cheddar2 (42% moisture, pH 5.3, 2.0% NaCl, aw 0.955)
Cheddar1 (42% moisture, pH 5.6, 1.8% NaCl, aw 0.945)
Cheddar3 (40% moisture, pH 5.3, 2.5% NaCl, aw 0.935)
All formulations contain 0.2% potassium sorbate
24
Formulation Safety for Process Cheese Spread
FRI Model (aka Tanaka Model) Safety factors for control of Clostridium botulinum in shelf-stable process cheeses
Moisture
pH
Sodium chloride
Sodium phosphate
Not modeled
Aw
lactate
Tanaka et al, 1986 JFP
Formulation safety for non-standard process
cheese: sorbic acid and Clostridium botulinum
25
Note: 2017 model validation in
progress
Probability of failure for 2017 model
set to 0.001
Use for guidance only for product
developers
%Predicted time to toxicity (weeks)
moisture pH NaCl DSPtotal salts fat
sorbic acid FRI 2017 FRI 1986
54.0 5.8 2.0 1.7 3.70 22 0 7 7
54.0 5.8 2.0 1.7 3.70 22 0.1 36 7
54.0 5.8 2.0 1.7 3.70 22 0.15 80 7
54.0 5.8 2.0 1.7 3.70 22 0.2 179 7
Cultured celery, pH, product matrix
Refrigerated foods:
Proteolytic C. botulinum
Matrix pH Treatment Storage Temp
°F
Toxicity (wk)
Dijon Pork 6.0 Control 59 5
68 1
Cultured Celery
(80 ppm NO2)
59 >8
68 1
Cauliflower-
potatoes
5.5 Control 59 7
68 1
Cultured Celery(60 ppm NO2)
59 >8
Golden et al., 2017, JFP, in press 68 2
Received “non-prot botulinum cook” 90°C, 10 min
Tested for toxin weekly
No toxicity for pH <6.0 during storage for 8 weeks at 50°F
Dry vinegar, Fruit-Spice Blend, pH
Uncured Chicken: C. botulinum Time to toxin production
pH 6.15 pH 6.40
Temperature
°FControl
0.5% Dry Vinegar
+ 0.6% Fruit-
Spice-Vinegar
blend
Control
0.5% Dry Vinegar
+ 0.6% Fruit-
Spice-Vinegar
blend
77 2 d 4 d 2 d 3 d
55 1 mo 1 mo 1 mo 1 mo
45 >6 mo >6 mo 1 mo 3 mo
Summary
• Formulation alone will not guarantee safety
• Heat or alternative pasteurization
• Proper sanitation
• Part of the well designed food safety system
• Factors to consider• Storage temperature distribution, at retail, by consumers
• Water activity, pH/total acidity
• Synthetic and clean label antimicrobials
• Requires that manufacturing specifications are met
• Success also depends on education and
cooperation of consumer for safe food handling
Acknowledgements
• FRI Applied Food Safety Lab
• Meat and Muscle Biology Lab
• WI Center for Dairy Research
• Funding • University of Wisconsin Foundation
• North American Meat Institute Foundation
• USDA
• Dairy Management Inc.
• FRI Summer Scholar Program
• International Dairy Foods Association
• Wisconsin Association of Meat Processors
• Industry support
FSPCA PCQI Training (Human Foods)
• UW-Madison August 22-24, 2017
• 2.5 day standard curriculum
• Blended Course
• Part 1 online
• Part 2 instructor-led
• Other courses held around US, several times per month
• See https://fspca.force.com for registration links
• Upcoming classes in WI
• Cherney Microbiology
• Covance
• NSF
• Safe Food Resources
